Abstract
BackgroundThe downregulation of microRNA (miR)-451a has been reported in bladder cancer (BCa) tissues. Herein, we elucidated the role of miR-451a in BCa with the involvement of DNA methyltransferase 3B (DNMT3B).MethodsWe first screened the differentially expressed miRNAs from the serum of 12 BCa patients and 10 healthy controls in the BCa database GSE113486. Subsequently, we detected miR-451a expression and CpG island methylation of the promoter in BCa cells T24 and 5637 with DNMT3B knockdown. The downstream mRNAs of miR-451a were predicted by bioinformatics and KEGG enrichment analysis. Afterwards, the expression patterns of DNMT3B, miR-451a and erythropoietin-producing hepatocellular receptor tyrosine kinase class A2 (EPHA2) were altered in BCa cells to test the ability of cell proliferation, apoptosis, migration as well as invasion. Finally, the effect of miR-451a and DNMT3B was evaluated in vivo.ResultsmiR-451a was significantly reduced in serum of BCa patients and cell lines. Moreover, the expression of DNMT3B in BCa cells was significantly increased, thus promoting methylation of the miR-451a promoter, resulting in miR-451a inhibition. Additionally, we found that miR-451a targeted and negatively regulated EPHA2, while EPHA2 could activate the PI3K/AKT signaling, driving BCa cell growth and metastasis.ConclusionsOur study proposed and demonstrated that miR-451a downregulation mediated by DNMT3B is critical for proliferation, migration, and invasion of BCa, which may be beneficial for developing more effective therapies against BCa.
Highlights
The downregulation of microRNA-451a has been reported in bladder cancer (BCa) tissues
The antibodies used in the experiments were DNA methyltransferase 3B (DNMT3B) (#72335, Cell Signaling Technologies (CST), Beverly, MA, USA), phos-PI3K (#ab182651, Abcam, Cambridge, UK), phosAKT1 (#44-621G, Thermo Fisher Scientific Inc., Waltham, MA, USA), erythropoietin-producing hepatocellular receptor tyrosine kinase class A2 (EPHA2) (#37–4400, Thermo Fisher), KI67 (#9027, CST), glyceraldehyde-3-phosphate dehydrogenase (GAPDH, #ab181602, Abcam), horseradish peroxidase (HRP)-labeled goat anti-mouse (#G-21040, Thermo Fisher) or goat anti-rabbit IgG (#G-21234, Thermo Fisher)
Terminal deoxynucleotidyl transferase (TdT)-mediated dUTPbiotin nick end labeling (TUNEL) kit was purchased from Promega (#G3250, Madison, WI, USA). 5-Ethynyl-2′-deoxyuridine (EdU) cell proliferation detection kit was obtained from Solarbio (#CA1170, Beijing, China)
Summary
The downregulation of microRNA (miR)-451a has been reported in bladder cancer (BCa) tissues. We elucidated the role of miR-451a in BCa with the involvement of DNA methyltransferase 3B (DNMT3B). To significantly halt the course of the malignancy, improvements should be achieved in the diagnosis and treatment of metastatic BCa. MicroRNAs (miRNAs) are endogenous ~ 23 nt RNA transcripts that play vital gene-regulatory roles in animals. Deep-sequencing-based BCa signatures conducted by Matsushita et al suggested that clustered miRNAs, including miR-451a were downregulated in BCa tissues [6], with their specific role in BCa undetermined. It has been previously reported that upregulated heterochromatin protein 1γ in prostate cancer cells repressed the expression pattern of miR-451a by promoting H3K9 methylation at the promoter region of miR451a [8]. This study may provide new and promising therapeutic targets for BCa treatment
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